Search Results (196)

Gatifloxacin (GTFX), a fourth-generation fluoroquinolone, causes metabolic disturbances in mammals, but its hepatotoxic mechanisms in aquatic vertebrates remain unclear. This study investigated whether GTFX induces liver injury in zebrafish larvae through oxidative stress or alternative pathways. Larvae at 3 days post-fertilization were exposed to 0.2–2.3 mg/mL GTFX for 48 h. Liver morphology, histopathology, intracellular reactive oxygen species (ROS), and expression of lipid metabolism (pparg) and xenobiotic biotransformation genes (cyp1a, cyp1b1) were assessed. GTFX exposure caused concentration-dependent reductions in liver area, increased hepatic opacity, delayed yolk sac absorption, and hepatocyte swelling with cytoplasmic vacuolization. Despite these structural changes, ROS levels did not differ significantly from those of controls. In contrast, transcriptional analysis revealed significant upregulation of pparg, cyp1a, and cyp1b1, indicating disrupted lipid homeostasis and enhanced detoxification responses. Acute high-dose GTFX exposure induced a metabolism-associated hepatotoxic response in zebrafish larvae, which occurred without a statistically significant change in bulk ROS levels. Together, these findings offer mechanistic insight into fluoroquinolone-associated liver injury. Full article

Metabolic reprogramming is closely linked to tumor proliferation, invasion, and immune escape. Despite its central role in amino acid metabolism, the regulatory mechanisms of asparagine metabolism in hepatocellular carcinoma (HCC) progression remain poorly characterized. Rather than focusing on canonical metabolic genes, prognostic markers were identified from co-expression modules associated with asparagine metabolism signatures. Using the TCGA database and asparagine metabolism-related gene sets, a prognostic risk-scoring model was developed through differential expression analysis, univariate Cox regression, and the LASSO algorithm and externally validated with the GEO dataset (GSE14620). Survival analysis, ROC curve evaluation, nomogram construction, scRNA-seq, GSEA, and drug sensitivity analysis were performed to systematically delineate the molecular mechanisms by which asparagine metabolism drives HCC progression. A three-gene signature comprising BOP1, SAC3D1, and PDE2A effectively stratified patients into high- and low-risk groups. High-risk patients exhibited markedly poorer overall survival, enrichment in tumor proliferation-associated pathways, increased tumor purity, reduced immune cell infiltration, and a substantially higher TP53 mutation rate (38% vs. 13%). In contrast, the low-risk group showed enrichment in pathways linked to hepatoblastoma suppression and liver function, alongside improved predicted response to immunotherapy. Single-cell analysis identified NK cells and endothelial cells as central mediators of asparagine metabolism-driven HCC progression, with BOP1, SAC3D1, and PDE2A displaying dynamic expression patterns during differentiation. Furthermore, the high-risk group was predicted to be more sensitive to chemotherapeutics such as cyclophosphamide and 5-fluorouracil. These findings highlight a potential interplay between nitrogen metabolism and asparagine metabolism in HCC and suggest mechanisms by which these pathways may influence NK cell and endothelial cell function to promote disease progression. This study establishes a novel prognostic model and identifies potential chemotherapeutic vulnerabilities in high-risk patients, warranting further experimental and clinical validation. Full article

Background/Objectives: Triple-negative breast cancer (TNBC) is frequently characterized by notably elevated Ki-67 expression, a hallmark of uncontrolled rapid cell-cycle progression. However, the underlying mechanisms remain unclear, leading to limited therapeutic options. Methods: In this study, hub gene was identified through integrated bioinformatic analysis of public datasets (TCGA-BRCA and METABRIC). Subsequent functional validation was performed both in vitro and in vivo using siRNA-mediated knockdown and small-molecule inhibitors. Phenotypic effects—including cell viability, cell cycle distribution, DNA synthesis, and clonogenic survival—were comprehensively assessed using MTT assays, flow cytometry, EdU, and colony formation assays. Protein-level changes were confirmed by Western blotting and immunohistochemistry (IHC). To dissect the transcriptional regulation of the key hub gene TTK, we first predicted potential upstream transcription factors using the JASPAR database; binding specificity was then validated through in silico motif analysis, luciferase reporter assays, and chromatin immunoprecipitation followed by quantitative PCR (ChIP-qPCR). Results: The mitotic kinase TTK is significantly overexpressed in TNBC compared with non-TNBC breast cancers. Notably, TTK overexpression exhibited a strong positive correlation with elevated Ki-67 indices and reduced overall survival in TNBC patients. Functional validation demonstrated that pharmacological or genetic inhibition of TTK effectively induced G2/M cell-cycle arrest and potently suppressed TNBC proliferation in both in vitro cell cultures and in vivo xenograft models. Mechanistically, TTK overexpression stems from enhanced transcriptional initiation driven by the transcription factor NFYA binding to the CCAAT box in the TTK promoter—an interaction newly identified here. Concurrently, TTK blockade disrupted spindle assembly checkpoint (SAC) signaling via BUB1B/MAD1L1 downregulation, triggering mitotic arrest and catastrophe. Conclusions: Collectively, these findings establish TTK as a key cell-cycle regulator driving TNBC proliferation. More importantly, targeting mitotic control through TTK inhibition represents an efficient strategy to impede the aberrantly fast cell cycle progression in TNBC. Full article

The rising global demand for lithium has led to substantial accumulation of lithium slag, a by-product of lithium carbonate production and a potential environmental contaminant. Leachates from this material contain various metal elements and may pose risks to ecosystems and organismal health. However, research on its neurotoxicity and underlying mechanisms remains limited. In this study, zebrafish embryos at 6 h post-fertilisation were exposed to varying concentrations of lithium slag leachate for 7 days. The leachate contained multiple metal ions (Li, Fe, Mn, Ni, Zn, As, Cr, Cu, Hg, Cd, Pb, etc.). Following exposure, significant metal accumulation was observed in larvae, accompanied by developmental malformations (yolk sac oedema, cardiac haemorrhage, and uninflated swim bladders). Behavioural assessment revealed reduced swimming distance and velocity, along with disrupted circadian rhythms. Biochemical analyses showed elevated Reactive oxygen species (ROS), Superoxide dismutase (SOD), Catalase (CAT), and Malondialdehyde (MDA), alongside decreased Glutathione (GSH), indicating oxidative stress. Transcriptomic analysis confirmed downregulation of core circadian genes. Neurotransmitter assays revealed decreased acetylcholine (Ach), noradrenaline (NE), and dopamine (DA), with increased gamma-aminobutyric acid (GABA) and serotonin (5-HT). These findings demonstrate that lithium slag leachate induces oxidative stress, circadian disruption, and neurobehavioural toxicity in zebrafish, providing important evidence for environmental risk assessment. Full article

CRISPR screens are a powerful functional genomics approach for identifying genes that confer sensitivity and resistance to anti-cancer therapies. CFI-402257 (luvixasertib, 2257) is a small molecule inhibitor of threonine tyrosine kinase (TTK), a promising therapeutic target in genomically unstable cancers due to its critical role in establishing the spindle assembly checkpoint (SAC) during mitosis. To inform its ongoing development and evaluation in clinical trials, we sought to use CRISPR activation (i.e., gain of function) screens to identify cellular mechanisms of resistance to 2257 in models of triple-negative breast cancer (TNBC). In vitro screens conducted in two TNBC cell lines nominated ABCG2 as the top resistance-conferring gene in both models. Validation studies assessing clonogenic survival and apoptosis confirmed that ABCG2 overexpression enhanced TNBC resistance to 2257 in vitro, while knockdown enhanced sensitivity. These findings suggest that 2257 is a substrate of ABCG2’s drug efflux activity. However, overexpression of ABCG2 failed to confer resistance to 2257 in TNBC xenografts grown in mice and treated with a moderately active dose and schedule. Our results highlight the potential impact of drug transporters in in vitro CRISPR screens and the importance of confirming the relevance of drug response mechanisms identified in cultured cells using in vivo models that recapitulate drug pharmacokinetics and pharmacodynamics. Full article

Serrated adenocarcinoma (SAC) represents a molecularly heterogeneous subtype of colorectal carcinoma (CRC) linked to the serrated pathway. It is aimed to clarify the molecular mechanisms underlying SAC development. Digital slides from The Cancer Genome Atlas (TCGA) colorectal adenocarcinoma Firehose Legacy dataset (632 cases) were reviewed, and cases were classified as SAC, partial-SAC, or classical CRC. Genomic alterations, mRNA expression, and DNA hypermethylation were compared using cBioPortal. Enrichment analyses were performed via WebGestalt, and protein–protein interaction (PPI) networks with hub genes were identified using STRING and Cytoscape. Statistical significance was defined as p < 0.05 and q < 0.05. The results revealed that the groups showed significant differences in the expression of 327 genomic alterations, 20 mRNAs, and 21 methylated genes (p < 0.0001, q < 0.0001). Hub genes were PSMC1, FLT3LG, SNW1, H3C2, H1-2, H2BC14, H1-5, RPS16, SUPT5H, and MYOD1. The pathways associated with differently expressed genes were the following: cell structure and morphology (phagocytic vesicle, microvillus, endocytosis, and immobile cilium), protein kinase activity (particularly MAPK), and immunological mechanisms. The hub genes act as molecular bridges connecting the observed genomic and epigenetic variations, particularly driving chromatin-related regulation and MAPK signaling pathways. In particular, PSMC1, SNW1, H3C2, H1-2, and H2BC14 genes offer promising molecular targets for future therapeutic approaches in SACs. Full article

The granulation of pomelo (Citrus maxima) juice sacs severely compromises fruit quality and is closely associated with lignin accumulation, a process catalyzed by peroxidases (PODs). Analysis of ‘Sanhong’ pomelo juice sacs collected 175–215 days after flowering revealed that bound peroxidase (BPOD) activity paralleled changes in lignin content, suggesting a potential role for BPOD in lignin biosynthesis. A total of 71 CmPOD genes were identified in the pomelo genome through integrated HMMER and BLAST analyses. Among them, CmPOD52 was selected for functional characterization based on its alkaline peroxidase properties, absence of a CE domain, predicted extracellular localization, and gradually increasing expression pattern revealed by RT-qPCR. Its transient overexpression in ‘Sanhong’ pomelo juice sacs for 36 h increased BPOD activity 2.06-fold (p < 0.01) compared to the empty vector control, indicating that CmPOD52 may be a BPOD gene. The recombinant CmPOD52 protein was expressed in a prokaryotic system, purified, and used in enzymatic assays with sinapyl alcohol as the substrate. The recombinant CmPOD52 protein, assayed at 272 nm with controls (substrate-only blank and heat-inactivated protein), showed an activity of 13.67 ± 0.9 U. The experimental group showed new products, identified by mass spectrometry as sinapyl alcohol dimers, thus suggesting that the recombinant protein catalyzes the dehydrogenation and polymerization of sinapyl alcohol monomers. This study identified CmPOD52, a gene potentially involved in lignin polymerization in pomelo juice sacs, offering a key candidate for further in vivo validation. Full article

Mammalian placentation represents one of the most striking evolutionary innovations among vertebrates, and accumulating evidence indicates that virus-derived genes—particularly the metavirus-derived PEG10 and PEG11/RTL1—have played indispensable but distinct roles: PEG10 in the emergence of therian viviparity and PEG11/RTL1 in the subsequent differentiation between marsupial and eutherian placental types. Notably, the metavirus-derived SIRH/RTL gene group, which includes PEG10 and PEG11/RTL1, exhibits unique and diverse functions not only in placenta development but also within microglia of the brain. Because microglia originate from yolk sac progenitors, these findings suggest that extraembryonic tissues such as the placenta and yolk sac provided permissive environments that enabled the retention, expression and functional domestication of virus-derived sequences. Once the placenta itself was established through viral gene integration, it may in turn have acted as a powerful driver of eutherian evolution through recurrent acquisition and co-option of additional virus-derived genes—a process we refer to as “placenta-driven evolution.” This perspective offers a unified framework in which viral gene acquisition is viewed as a key driver of genomic innovation, tightly intertwined with the emergence of viviparity, subsequent divergence at the marsupial–eutherian split, and continued diversification of placental structure and function across eutherian lineages. Full article

Background and Clinical significance: Cowden syndrome is an autosomal dominant disorder caused by germline loss-of-function mutations in the PTEN tumor suppressor gene. It is characterized by multiple hamartomas and an increased lifetime risk of malignancies affecting the breast, thyroid, endometrium, and gastrointestinal (GI) tract. Pediatric presentations may include macrocephaly, scrotal tongue, and intellectual disability. Gastrointestinal involvement is frequent, with juvenile-like hamartomatous polyps occurring in at least half of patients and distributed throughout the GI tract, posing a risk for malignant transformation. Early diagnosis and surveillance are crucial for improving patient outcomes. Case Presentation: We report a case of a 10-year-old Caucasian female with Cowden syndrome, with a history of a malignant germ cell tumor of the ovary consisting of a yolk sac tumor and low-grade immature teratoma diagnosed at age six, and thyroidectomy at age nine. The patient has mild intellectual disability. Routine radiological surveillance revealed a right colon intraluminal mass, prompting referral for pediatric gastroenterology evaluation. Endoscopy identified multiple polyps throughout the colon, stomach, and small intestine. Polypectomy of larger lesions was performed, and histopathology confirmed juvenile-like hamartomatous polyps without dysplasia or malignancy. This case highlights the necessity of comprehensive gastrointestinal evaluation in pediatric Cowden syndrome patients. Endoscopic surveillance is essential for early detection and management of polyps. Conclusions: Given the multisystem involvement and elevated cancer risk associated with PTEN mutations, a multidisciplinary approach that includes genetic counseling, dermatologic evaluation, and ongoing oncologic monitoring is recommended. Increased awareness of gastrointestinal manifestations enables timely intervention and may reduce morbidity and mortality in this high-risk population. Full article

Erythro-myeloid progenitors (EMPs) originate from the haemogenic endothelium in the yolk sac via an endothelial-to-haematopoietic transition (EHT) to generate blood and immune cells that support embryo development. Yet, the transitory nature of EHT and the limited availability of molecular markers have constrained our understanding of the origin, identity, and differentiation dynamics of EMPs. Here, we have refined the annotation of yolk sac haemato-vascular populations in publicly available single-cell RNA sequencing (scRNAseq) datasets from mouse embryos to identify novel molecular markers of haemogenic endothelium and EMPs. By sub-clustering key cell populations followed by pseudotime analysis, we refined cluster annotations and then reconstructed differentiation trajectories. Subsequent differential gene expression analysis between clusters identified novel cell surface markers for haemogenic endothelial cells (Fxyd5 and Scarf1) and EMPs (Fcer1g, Tyrobp, and Mctp1). Further, we have identified candidate signalling and metabolic pathways that may regulate yolk sac haematopoietic emergence and differentiation. The specificity of FXYD5, SCARF1, and FCER1G for haemogenic endothelium and EMPs was validated by immunostaining of the mouse yolk sac. These insights into the transcriptional dynamics in the yolk sac should support future investigation of EHT and haematopoietic differentiation during early mammalian development. Full article

Background: The inner ear hosts several macrophage populations. Endolymphatic sac macrophages can phagocytose otoconia, and spiral limbus macrophages express genes for fluid shear stress sensing and bone remodeling. Obstruction of endolymph flow by saccular otoconia could be linked to endolymphatic hydrops. Since macrophages are strongly affected by inflammatory status, a role for them in otolith removal could provide a link between inflammation and hydrops. However, the distribution of macrophages around the reuniting duct (RD) and endolymphatic duct (ED), which are narrow structures likely prone to blockage, remains unexplored. Methods: We performed tissue clearing and light-sheet imaging on rat temporal bones. Autofluorescence and immunolabeling for collagen IV, smooth muscle actin, and Iba1 were used to visualize inner ear structures, blood vessels, and macrophages. Results: The connective tissue layer underlying the RD extended from the cochlear spiral limbus. The RD and spiral limbus hosted a continuous microvascular network and macrophage population, comprising both ameboid and ramified cells; macrophages also surrounded the underlying vestibulocochlear artery (VCA). A separate macrophage population, continuous with that of the saccular connective tissue, was found around the endolymphatic sinus and utriculo–endolymphatic (Bast’s) valve; macrophage patterns changed in the vestibular aqueduct and endolymphatic sac. Conclusions: Macrophages are observed in positions consistent with potential roles in sensing luminal changes and in the clearance of obstructive material from the RD and ED; functional confirmation will require targeted experiments. Full article

Von Hippel–Lindau disease-associated endolymphatic sac tumors (VHL-associated ELSTs) present diagnostic challenges due to their rarity and nonspecific symptoms. This study describes clinical, pathological and genotypic features to guide treatment. We retrospectively analyzed seven patients with VHL-associated ELSTs. The mean age of otologic symptom [hearing loss (100%) and facial nerve paralysis (85.71%)] onset was 22.43 ± 8.68 years (range: 10–33). Surgical management included trans-labyrinthine and subtotal temporal bone resection approaches. Among three patients with severe preoperative facial nerve dysfunction, two underwent great auricular nerve grafting improved to House–Brackmann grade IV, while one receiving hypoglossal–facial nerve anastomosis reached grade V. Genetic testing identified pathogenic VHL gene missense mutations in three patients. Two female patients demonstrated disease progression during pregnancy. Literature analysis revealed exon-specific patterns: Exon 1 mutations correlated with cerebellar/spinal hemangioblastomas in female patients, while Exon 3 mutations were associated with multisystem tumors. These findings support that VHL-associated ELSTs manifest early with otologic symptoms and demonstrate exon-specific phenotypic patterns. Optimal management requires complete surgical resection, genetic diagnosis, and a multidisciplinary approach to address these complex tumors and achieve favorable outcomes. Full article

The dedifferentiation of smooth muscle cells (SMCs) is the main cause of atherosclerosis and vascular calcification. This study integrated the gene expression data of multiple microarrays to identify relevant marker molecules. A total of 72 Gene Expression Omnibus (GEO) samples (GSM) were collected from 10 gene expression data series (GSE) and divided into five groups: non-SMC, SMC, atherosclerotic SMC (SMC-ath), calcified SMC (SMC-calc), and treated SMC (SMC-t). The SMC-t group included synthetic SMCs that had undergone treatment to inhibit proliferation, migration, or inflammation. The gene expression data were merged, normalized, and batch effects were removed before differential gene expression (DGE) analysis was performed via linear models for microarray data (limma) and statistical analysis of metagenomic profiles (STAMPs). The genes with expressions that significantly differed were subsequently subjected to protein-protein interaction (PPI) and functional prediction analyses. In addition, the random forest method was used for classification. Twelve proteins that may be marker molecules for SMC differentiation and dedifferentiation were identified, namely, Proprotein convertase subtilisin/kexin type 1 (PCSK1), Transforming growth factor beta-induced (TGFBI), Complement C1s (C1S), Phosphomannomutase 1 (PMM1), Claudin 7 (CLDN7), Calcium binding and coiled-coil domain 2 (CALCOCO2), SAC3 domain-containing protein 1 (SAC3D1), Natriuretic peptide B (NPPB), Monoamine oxidase A (MAOA), Regulator of the Cell Cycle (RGCC), Alpha-crystallin B Chain (CRYAB), and Alcohol dehydrogenase 1B (ADH1B). Finally, their possible roles in SMCs are discussed. This study highlights the feasibility of bioinformatics analysis for studying SMC dedifferentiation. Full article

Levan and levan-type fructooligosaccharides (L-FOSs) are non-digestible fructans with prebiotic properties that promote gut microbiota growth. This study presents the first genomic analysis of a Bacillus velezensis HL25 strain with high fructan-producing efficiency, revealing genes involved in sucrose utilization and fructan biosynthesis. A putative levansucrase operon was identified in the HL25 genome, consisting of the sacB levansucrase gene classified in GH68 subfamily 1 and the following three GH32 genes: endo-levanase (lev), β-fructofuranosidase (ffase), and sucrose-6-phosphate hydrolase (scrB). Remarkably, sugars involved in levan biosynthesis are proposed to be transported through three distinct systems: a multiple-component ABC sugar transporter, a glucose/H⁺ symporter, and glucose- and fructose-specific phosphotransferase systems (PTS). Subsequently, recombinant HL25SacB levansucrase exhibited optimal activity at 40 °C and pH 5.0 in 50 mM sodium acetate buffer. The enzyme demonstrates high specificity in converting sucrose into a mixture of short-chain FOSs (DP 2–4) and levan, achieving a 62.5% conversion rate at 30 °C with 200 g/L sucrose over 24 h. These findings demonstrate the potential of this B. velezensis HL25 strain as an efficient whole-cell biocatalyst and highlight the applicability of the recombinant HL25SacB enzyme as a promising tool for sustainable production of FOSs and levan. Full article

Scale adhesion strength is a key trait in aquaculture, directly influencing disease resistance, survival, and commercial value. The Dongting crucian carp (Carassius auratus indigentiaus, hereafter CaDT) is valued for its rapid growth and superior flesh quality but is characterized by loosely attached scales. In this study, we investigated the morphological and molecular basis underlying scale adhesion by comparing CaDT with the tight-scaled allogynogenetic gibel carp, Zhongke No. 3 (Carassius auratus gibelio, hereafter CaGB). Morphological analysis revealed a significantly lower scale-embedding ratio in CaDT compared to CaGB. To unravel the molecular mechanisms underpinning these phenotypic differences, a comparative transcriptomic analysis was conducted on scale sac, skin, and muscle tissues in CaDT and CaGB. In CaGB, gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses of differentially expressed genes (DEGs) in the critical scale sac tissue showed a significant upregulation of genes involved in ribosomal pathways. Specifically, key epithelial differentiation markers, including keratin 13 (krt13), keratin 15 (krt15), and metabolic genes, enolase 3-like (eno3l), and phosphoglycerate mutase 2 (pgam2) were significantly down-regulated in CaDT, which suggests a compromised epithelial cell differentiation capacity and reduced energetic and biosynthetic activity. Quantitative PCR (qPCR) validation across three tissues showed high concordance with the RNA-seq results, thereby confirming the reliability of the transcriptomic data. The results offer insight into the molecular basis for understanding scale adhesion traits, and provide valuable insights for selective breeding strategies to improve scale retention in aquaculture species. Full article